ABSTRACT
BACKGROUND:Piezo1,a mechanosensitive protein,is tightly connected to osteogenic differentiation,and it has been demonstrated that TAZ has a role in regulating osteogenic differentiation.It is unclear whether TAZ participates in the regulation of osteogenic differentiation of human bone marrow mesenchymal stem cells by Piezo1,so it is crucial to investigate its unique mechanism to prevent osteonecrosis of the femoral head. OBJECTIVE:To elucidate what function Piezo1 plays in osteogenic differentiation and TAZ expression in human bone marrow mesenchymal stem cells. METHODS:The siRNA targeting Piezo1 was constructed and transfected into 293T cells.The silencing efficiency was detected by RT-qPCR.The selected Piezo1-Home-2337 was packaged according to the silencing efficiency,and its optimal multiplicity of infection value was assayed by immunofluorescence staining.The packaged Piezo1 silencing recombinant lentivirus was transfected into human bone marrow mesenchymal stem cells,and its silencing effect was detected by RT-qPCR and western blot assay.Alizarin red staining,alkaline phosphatase activity analysis,immunofluorescence staining,RT-qPCR and western blot assay were utilized to analyze the effect of silencing Piezo1 on the osteogenic differentiation of human bone marrow mesenchymal stem cells. RESULTS AND CONCLUSION:(1)The mRNA and protein levels of Piezo1 in human bone marrow mesenchymal stem cells transfected by si-Piezo1 were decreased significantly,with a statistically significant difference compared with normal and negative control groups.(2)The alkaline phosphatase activity in the si-Piezo1 group was much lower and the calcium deposition in the si-Piezo1 group was significantly reduced compared with the negative control group.(3)The mRNA levels of osteogenesis-related genes including Runt-related transcription factor 2(Runx2),osteopontin(OPN),distal-less homeobox 5(DLX5),osteocalcin,β-catenin and Tafazzin(TAZ)in the si-Piezo1 group were significantly decreased compared with the negative control group.Afterward,the expression levels of TAZ and β-catenin protein in the si-Piezo1 group were down-regulated significantly compared with the negative control group,whereas the expression levels of p-TAZ and p-β-catenin protein in the si-Piezo1 group had the opposite condition.(4)The results of immunofluorescence staining showed that the expression of TAZ and β-catenin in human bone marrow mesenchymal stem cells in the si-Piezo1 group was less compared with the negative control group.(5)These findings indicate that Piezo1 can promote the osteogenic differentiation of human bone marrow mesenchymal stem cells.The osteogenic ability of human bone marrow mesenchymal stem cells is significantly reduced after silencing Piezo1,and the expression of TAZ is also reduced.
ABSTRACT
OBJECTIVE@#To establish finite element models of different preserved angles of osteonecrosis of the femoral head (ONFH) for the biomechanical analysis, and to provide mechanical evidence for predicting the risk of ONFH collapse with anterior preserved angle (APA) and lateral preserved angle (LPA).@*METHODS@#A healthy adult was selected as the study object, and the CT data of the left femoral head was acquired and imported into Mimics 21.0 software to reconstruct a complete proximal femur model and construct 3 models of necrotic area with equal volume and different morphology, all models were imported into Solidworks 2022 software to construct 21 finite element models of ONFH with LPA of 45°, 50°, 55°, 60°, 65°, 70°, and 75° when APA was 45°, respectively, and 21 finite element models of ONFH with APA of 45°, 50°, 55°, 60°, 65°, 70°, 75° when LPA was 45°, respectively. According to the physiological load condition of the femoral head, the distal femur was completely fixed, and a force with an angle of 25°, downward direction, and a magnitude of 3.5 times the subject's body mass was applied to the weight-bearing area of the femoral head surface. The maximum Von Mises stress of the surface of the femoral head and the necrotic area and the maximum displacement of the weight-bearing area of the femoral head were calculated and observed by Abaqus 2021 software.@*RESULTS@#The finite element models of ONFH were basically consistent with biomechanics of ONFH. Under the same loading condition, there was stress concentration around the necrotic area in the 42 ONFH models with different preserved angles composed of 3 necrotic areas with equal volume and different morphology. When APA was 60°, the maximum Von Mises stress of the surface of the femoral head and the necrotic area and the maximum displacement of the weight-bearing area of the femoral head of the ONFH models with LPA<60° were significantly higher than those of the models with LPA≥60° ( P<0.05); there was no significant difference in each index among the ONFH models with LPA≥60° ( P>0.05). When LPA was 60°, each index of the ONFH models with APA<60° were significantly higher than those of the models with APA≥60° ( P<0.05); there was no significant difference in each index among the ONFH models with APA≥60° ( P>0.05).@*CONCLUSION@#From the perspective of biomechanics, when a preserved angle of ONFH is less than its critical value, the stress concentration phenomenon in the femoral head is more pronounced, suggesting that the necrotic femoral head may have a higher risk of collapse in this state.